D. Methyl-CpG binding protein 2 (Mecp2/MECP2) is thought to selectively bind methyl-CpG dinucleotides in the mammalian genome and to function as a transcriptional represser in vivo by interacting with SinSA and recruiting histone deaceylase (Hdac). Mutations in MECP2 are associated with Rett syndrome, an X-linked neurodevelopmental disorder. We recently found that a maternally expressed imprinted gene, Dlx5, is a direct target gene for Mecp2 in mouse brain and in lympholastoid cells from individuals with Rett syndrome. DLX5 showed a loss of imprinting. As a result, Dlx5 expression is increased 2-fold when Mecp2 is absent or mutated. We also reported that Mecp2 regulates genes by forming a chromatin loop specifically in transcriptionally silent chromatin. DLX5 regulates the production of (-aminobutyric acid (GABA). Using transgenic mice expressing increased levels of DLX5, we propose to study whether Dlx5/DLX5 dysregulation per se is sufficient to cause some of the neurological phenotypes of Rett syndrome. We will investigate this question by systematic behavioral analyses of the DLX5 transgenic mice. We will study expression of Dlx5 and Gad, an enzyme which synthesizes GABA, as well as GABA itself, in various subregions of the brains of wild-type, Mecp2-null, and DLX5 transgenic mice (Specific Aim 1 and 2). In addition to Dlx5, we recently identified another imprinted gene, Peg3, to be dysregulated in the hippocampus of the Mecp2-null brain. We will study whether Peg3 is also a direct target gene for Mecp2, and if it is, we will determine how Mecp2 regulates expression of this gene (Specific Aim 3). Because Peg3 is known to be linked to maternal behaviors, we will evaluate the maternal behaviors of Mecp2 (+/-) female mice expressing increased levels of Peg3 in their brain. Furthermore, we will evaluate Peg3 overexpression in myogenesis by the in vitro differentiation cell culture system (Specific Aim 4). This is because Peg3 is specifically and abundantly expressed in the brain and skeletal muscle in adult mice. Our study will provide information about the molecular mechanisms underlying Mecp2 activity and how MECP2 mutations might cause Rett syndrome.